The light-harvesting protein Lhca2 is one of the four main and highly conserved types of chlorophyll a/b-binding proteins (Lhca1-4) of the light harvesting antenna (LHCI) of plant photosystem I. Lhca2 is imported as a precursor from the cytosol into the chloroplast. Upon integration in the thylakoid membrane Lhca2 forms a heterodimer (LHCI-680) with Lhca3 that associates with the PSI core close to PsaF and PsaK.A biochemical characterization of the plant LHCI antenna can be found in Klimmek et al. (2005) The structure of the higher plant light harvesting complex I: in vivo characterization and structural interdependence of the Lhca proteins. Biochemistry 44: 3065–3073.

Immunogen

BSA-conjugated synthetic peptide derived from the Lhca2 protein of Arabidopsis thaliana UniProt: Q9SYW8, Q8LCQ4, TAIR: At3g61470. This sequence is highly conserved in Lhca2 proteins of angiosperms (monocots and dicots) and gymnosperms as well as in At1g19150. This gene codes for the very low expressed Lhca6 protein which also has been denoted as Lhca2*1.

Host

Rabbit

Clonality

Polyclonal

Clone

Purity

Total IgG

Format

Lyophilized in PBS pH 7.4.

Quantity

0.5 mg

Reconstitution

For reconstitution add 100 µl of sterile water.

Storage

Store lyophilized/reconstituted at -20°C; once reconstituted make aliquots to avoid repeated freeze-thaw cycles. Please, remember to spin tubes briefly prior to opening them to avoid any losses that might occur from lyophilized material adhering to the cap or sides of the tubes.

1.0 µg of chlorophyll from mesophyll (M) and bundle sheath (BS) thylakoids of various C4 plants (Echinochloa crus-galli, Panicum miliaceum, Zea mays) extracted with 0.4 M sorbitol, 50 mM Hepes NaOH, pH 7.8, 10 mM NaCl, 5 mM MgCl2 and 2 mM EDTA. Samples were denatured with Laemmli buffer at 75 0C for 5 min and were separated on 12% SDS-PAGE and blotted 30 min to PVDF using wet transfer. Blot was blocked with 5% milk for 1h at room temperature (RT) with agitation. Blot was incubated in the primary antibody at a dilution of 1: 2000 overnight at 4°C with agitation in 1% milk in TBS-T. The antibody solution was decanted and the blot was washed 4 times for 5 min in TBS-T at RT with agitation. Blot was incubated in secondary antibody (anti-rabbit IgG horse radish peroxidase conjugated, from Agrisera, AS09 602, Lot 1702) diluted to 1:25 000 in 1 % milk in TBS-T for 1h at RT with agitation. The blot was washed 5 times for 5 min in TBS-T and 2 times for 5 min in TBS, and developed for 1 min with 1.25 mM luminol, 0.198 mM coumaric acid and 0.009% H2O2 in 0.1 M Tris- HCl, pH 8.5. Exposure time in ChemiDoc System was 15 seconds.

Courtesy of Dr. Wioleta Wasilewska, Warsaw University, Poland

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